Augmented cytokine production by tissue macrophages has been implicated in the pathogenesis of organ dysfunction, and may contribute to hepatic and pulmonary dysfunction after thermal injury. The mechanisms for altered cytokine production following thermal injury are poorly characterized. Inter- and intra-organ signaling mechanisms have been shown to regulate tissue macrophage cytokine production during inflammation. Hepatic secretory products, acting locally on the kupffer cell or distantly on the alveolar macrophage, may represent one such signaling pathway. In addition, autocrine mechanisms regulate macrophage inflammatory responses. We propose to investigate tissue macrophage cytokine responses following thermal injury and mechanisms which regulate these responses. The central hypothesis is that thermal injury augments cytokine production by tissue macrophages and that this response is regulated, in part, by hepatic secretory products or altered autocrine mechanisms. The specific aims for this proposal are: 1) characterize the effect of thermal injury on tissue macrophage TNF and IL-6 production; 2) determine whether thermal injury induces hepatocyte secretory products that regulate kupffer cell and alveolar macrophage responses; 3) determine the effect of thermal injury on autocrine mechanisms regulating cytokine production. A resuscitated, rodent model of thermal injury will be utilized for this project. Alveolar and hepatic macrophage TNF and IL-6 production will be determined at the levels of bioactivity, cell-associated protein, and mRNA after thermal injury alone and injury followed by lipopolysaccharide utilizing cytokine dependent cell lines, immunoprecipitation, and a nuclease protection assay, respectively. Changes with time and severity of thermal injury will be assessed. In vivo serum and tissue cytokine levels, tissue histology, and mortality will be performed to correlate with in vitro findings. The role of hepatic secretory products on cytokine production will be examined in supernatant transfer experiments. Supernatants of hepatocytes from thermally injured animals will be added to tissue macrophages and effects on cytokine production examined; subsequently any hepatocyte-derived regulatory signals will be biochemically characterized and proteins purified by preparative SDS-PAGE. The production of LPS- binding protein (LBP), a specific hepatocyte acute phase reactant, will be examined following thermal injury at the protein synthesis and mRNA levels. The autoregulatory role on TNF receptor expression and PGE2 release in tissue macrophage cytokine production after thermal injury will be investigated. TNF receptor expression will be determined in binding studies and receptor synthesis regulation determined at protein and mRNA levels. The role of macrophage desensitization to PGE2 as a mechanism of increased cytokine production will be examined by determining its release, induction of cAMP, and its effect on cytokine production after thermal injury. These results will define macrophage function contributing to organ dysfunction and contribute to developing therapeutic strategies to manipulate macrophage function after thermal injury. P50GM216819001 The Core, in addition to administrative and logistic functions, will furnish animal models and technology that augments each of the four projects, and will develop technology, useful in treatment application, both experimentally and clinically. MRI spectrometry will be used to define the bioenergetics, ion concentrations and intracellular pH of various organs, determination of blood flow with blood flow measurements by laser Doppler; and estimation of the redox potential by serial measurements of conjugated dienes and malondialdehyde and progressive changes in antioxidant activity by glutathione reductase. The Core will provide the wound model in pigs that will be used in the evaluation of local perturbations and for testing wound healing, including the efficacy of composite grafts developed in the core; altering proteinase activities (dipeptidyl peptidase etc.) and measured augmentation of wound vascularization (laser Doppler). Liposomal encapsulation of compounds useful for therapeutic interventions are being developed and the possible interaction of the micelle with phagocytic cells and cytokines explored.